胰岛素样生长因子1与转化生长因子β_2促进组织工程软骨构建的体外实验

Insulin-like growth factor 1 and transformation growth factor beta 2 promote engineered cartilage formation in vitro

背景:软骨组织的再生能力差,软骨组织工程能利用较少的细胞、支架材料和细胞因子对缺损进行修复。目的:观察胰岛素样生长因子1与转化生长因子β2联合应用对组织工程软骨形成的影响。方法:用酶消化法获取人软骨细胞,将培养的细胞以4×109L-1的细胞浓度接种在藻酸钙凝珠支架上,分别加入200μg/L胰岛素样生长因子1和(或)1μg/L转化生长因子β2进行立体培养。于培养的第3,5,7,9,11,13天,进行细胞计数,观察软骨细胞的增殖情况。培养2周后,进行大体形态观察和阿尔新蓝-过碘酸雪夫氏染色(AB-PAS)及抗Ⅱ型胶原免疫组织化学染色。结果与结论:细胞计数及免疫组织化学染色显示,胰岛素样生长因子1和转化生长因子β2均能促进软骨细胞增殖和软骨相关基质黏多糖及Ⅱ型胶原的分泌,其中胰岛素样生长因子1的作用主要体现在促细胞增殖方面,而转化生长因子β2的作用主要体现在促进软骨相关基质形成方面,二者联合应用具有促进组织工程软骨形成的协同作用。

BACKGROUND：Cartilage tissue has poor regenerative capacity.Cartilage tissue engineering can repair damaged cartilage tissue using fewer cells,scaffold materials and cytokines.OBJECTIVE：To investigate the effects of insulin-like growth factor（IGF-1） combined with transformation growth factor beta 2（TGF-β2） on engineered cartilage formation.METHODS：Human chondrocytes were isolated by enzyme digestion method and then were inoculated onto calcium alginate bead scaffold for culture in three-dimensional circumstance after addition of 200 μg/L IGF-1 and（or） 1 μg/L TGF-β2.At 3,5,7,9,11,and 13 days of culture,chondrocyte proliferation was tested by cell counting.After 2 weeks of culture,the fresh tissues were assessed by morphological observation,alcian blue-periodic acid sthiff staining,and anti typeⅡcollagen immunohistochemicel staining.RESULTS AND CONCLUSION：IGF-1 and TGF-β2 could stimulate chondrocytes to proliferate and excrete chondral matrix.IGF-1 mainly promoted the proliferation of chondrocytes,while TGF-β2 mainly promoted the formation of cartilage-related matrix.IGF-1 combined with TGF-β2 has synergistic effects on promoting engineered cartilage formation.